A determination of the sub-units of arachin by osmometry. Arachins A, B and A1

Abstract

Osmotic pressure determinations of dissociated arachins are a particularly suitable method for determination of the number of sub-units in tne protein, because they yield a number-average molecular weight. Arachin, in 8M-urea-0.1 M-sulfite, produced 12 sub-units from the form of molecular weight 345,000. When the urea concentration was varied the molecules became fully dissociated at 6M-urea-0.1 M-sulfite. Although sulfite was necessary to break disulfide bridges, concentrations greater than 0.1 M caused a reaggre-gation of the sub-units. Similar results were obtained in guanidine solutions. A new form of arachin has been discovered, A1, migrating more rapidly than arachin A. The N-terminal residues of arachin were reinvestigated on more highly purified samples: they were gly-cine, valine and (iso)leucine in the proportions 4:1:1:6. The 3 forms of arachin have the structure (B) [beta]4 [gamma][delta], (A) [alpha]2[beta]2 [gamma] [delta] and (A1) [alpha]4 [gamma] [delta], for the forms of molecular weight 170,000. Dissociation in 8M-urea produces some fragments, detected by gel electrophoresis, which appear to be dimers of the type [alpha]-S-S-[beta], [beta]-S-S-[beta], held together by disulfide bonds.